Dlk2 interacts with Syap1 to activate Akt signaling pathway during osteoclast formation

AUTHORS

Xinwei Chen, Xuzhuo Chen, Rui Chao, Yexin Wang, Yi Mao, Baoting Fan, Yaosheng Zhang, Weifeng Xu, An Qin & Shanyong Zhang

ABSTRACT

Excessive osteoclast formation and bone resorption are related to osteolytic diseases. Delta drosophila homolog-like 2 (Dlk2), a member of the epidermal growth factor (EGF)-like superfamily, reportedly regulates adipocyte differentiation, but its roles in bone homeostasis are unclear. In this study, we demonstrated that Dlk2 deletion in osteoclasts significantly inhibited osteoclast formation in vitro and contributed to a high-bone-mass phenotype in vivo. Importantly, Dlk2 was shown to interact with synapse-associated protein 1 (Syap1), which regulates Akt phosphorylation at Ser473. Dlk2 deletion inhibited Syap1-mediated activation of the AktSer473, ERK1/2 and p38 signaling cascades. Additionally, Dlk2 deficiency exhibits increased bone mass in ovariectomized mice. Our results reveal the important roles of the Dlk2-Syap1 signaling pathway in osteoclast differentiation and osteoclast-related bone disorders.

Gulp1 deficiency augments bone mass in male mice by affecting osteoclasts due to elevated 17β-estradiol levels

AUTHORS

Soon-Young Kim, Gun-Il Park, Seung-Yoon Park, Eun-Hye Lee, Hyuck Choi, Jeong-Tae Koh, Soyun Han, Man Ho Choi, Eui Kyun Park, In-San Kim, Jung-Eun Kim

ABSTRACT

The engulfment adaptor phosphotyrosine-binding domain containing 1 (GULP1) is an adaptor protein involved in the engulfment of apoptotic cells via phagocytosis. Gulp1 was first found to promote the phagocytosis of apoptotic cells by macrophages, and its role in various tissues, including neurons and ovaries, has been well studied. However, the expression and function of GULP1 in bone tissue are poorly understood. Consequently, to determine whether GULP1 plays a role in the regulation of bone remodeling in vitro and in vivo, we generated Gulp1 knockout (KO) mice. Gulp1 was expressed in bone tissue, mainly in osteoblasts, while its expression is very low in osteoclasts. Microcomputed tomography and histomorphometry analysis in 8-week-old male Gulp1 KO mice revealed a high bone mass in comparison with male wild-type (WT) mice. This was a result of decreased osteoclast differentiation and function in vivo and in vitro as confirmed by a reduced actin ring and microtubule formation in osteoclasts. Gas chromatography-mass spectrometry analysis further showed that both 17β-estradiol (E2) and 2-hydroxyestradiol levels, and the E2/testosterone metabolic ratio, reflecting aromatase activity, were also higher in the bone marrow of male Gulp1 KO mice than in male WT mice. Consistent with mass spectrometry analysis, aromatase enzymatic activity was significantly higher in the bone marrow of male Gulp1 KO mice. Altogether, our results suggest that GULP1 deficiency decreases the differentiation and function of osteoclasts themselves and increases sex steroid hormone-mediated inhibition of osteoclast differentiation and function, rather than affecting osteoblasts, resulting in a high bone mass in male mice. To the best of our knowledge, this is the first study to explore the direct and indirect roles of GULP1 in bone remodeling, providing new insights into its regulation.

A Zeb1/MtCK1 metabolic axis controls osteoclast activation and skeletal remodeling

AUTHORS

Lingxin Zhu, Yi Tang, Xiao-Yan Li, Samuel A Kerk, Costas A Lyssiotis, Wenqing Feng, Xiaoyue Sun, Geoffrey E Hespe, Zijun Wang, Marc P Stemmler, Simone Brabletz, Thomas Brabletz, Evan T Keller, Jun Ma, Jung-Sun Cho, Jingwen Yang, Stephen J Weiss

ABSTRACT

Osteoclasts are bone-resorbing polykaryons responsible for skeletal remodeling during health and disease. Coincident with their differentiation from myeloid precursors, osteoclasts undergo extensive transcriptional and metabolic reprogramming in order to acquire the cellular machinery necessary to demineralize bone and digest its interwoven extracellular matrix. While attempting to identify new regulatory molecules critical to bone resorption, we discovered that murine and human osteoclast differentiation is accompanied by the expression of Zeb1, a zinc-finger transcriptional repressor whose role in normal development is most frequently linked to the control of epithelial-mesenchymal programs. However, following targeting, we find that Zeb1 serves as an unexpected regulator of osteoclast energy metabolism. In vivo, Zeb1-null osteoclasts assume a hyperactivated state, markedly decreasing bone density due to excessive resorptive activity. Mechanistically, Zeb1 acts in a rheostat-like fashion to modulate murine and human osteoclast activity by transcriptionally repressing an ATP-buffering enzyme, mitochondrial creatine kinase 1 (MtCK1), thereby controlling the phosphocreatine energy shuttle and mitochondrial respiration. Together, these studies identify a novel Zeb1/MtCK1 axis that exerts metabolic control over bone resorption in vitro and in vivo.

Suppression of osteoclast multinucleation via a posttranscriptional regulation–based spatiotemporally selective delivery system

AUTHORS

Qingqing Wang, Haoli Wang, Huige Yan, Hongsen Tian, Yining Wang, Wei Yu, Zhanqiu Dai, Pengfei Chen, Zhaoming Liu, Ruikang Tang, Chao Jiang, Shunwu Fan, Xin Liu, Xianfeng Lin

ABSTRACT

Redundancy of multinucleated mature osteoclasts, which results from the excessive fusion of mononucleated preosteoclasts (pOCs), leads to osteolytic diseases such as osteoporosis. Unfortunately, the currently available clinical drugs completely inhibit osteoclasts, thus interfering with normal physiological bone turnover. pOC-specific regulation may be more suitable for maintaining bone homeostasis. Here, circBBS9, a previously unidentified circular RNA, was found to exert regulatory effects via the circBBS9/miR-423-3p/Traf6 axis in pOCs. To overcome the long-standing challenge of spatiotemporal RNA delivery to cells, we constructed biomimetic nanoparticles to achieve the pOC-specific targeted delivery of circBBS9. pOC membranes (POCMs) were extracted to camouflage cationic polymer for RNA interference with circBBS9 (POCM-NPs@siRNA/shRNAcircBBS9). POCM-NPs endowed the nanocarriers with improved stability, accurate pOC targeting, fusogenic uptake, and reactive oxygen species–responsive release. In summary, our findings may provide an alternative strategy for multinucleated cell–related diseases that involves restriction of mononucleated cell multinucleation through a spatiotemporally selective delivery system.

Role of chromatin modulator Dpy30 in osteoclast differentiation and function

AUTHORS

Yanfang Zhao, Xiaoxiao Hao, Zhaofei Li, Xu Feng, Jannet Katz, Suzanne M.Michalek, Hao Jiang, Ping Zhang

ABSTRACT

Osteoclasts are the principal bone resorption cells crucial for homeostatic bone remodeling and pathological bone destruction. Increasing data demonstrate a vital role of histone methylation in osteoclastogenesis. As an integral core subunit of H3K4 methyltransferases, Dpy30 is notal as a key chromatin regulator for cell growth and differentiation and stem cell fate determination, particularly in the hematopoietic system. However, its role in osteoclastogenesis is currently unknown. Herein, we generated Dpy30F/F; LysM-Cre+/+ mice, which deletes Dpy30 in myeloid cells, to characterize its involvement in osteoclast differentiation and function. Dpy30F/F; LysM-Cre+/+ mice showed increased bone mass, evident by impaired osteoclastogenesis and defective osteoclast activity, but no alteration of osteoblast numbers and bone formation. Additionally, our ex vivo analysis showed that the loss of Dpy30 significantly impedes osteoclast differentiation and suppresses osteoclast-related gene expression. Moreover, Dpy30 deficiency significantly decreased the enrichment of H3K4me3 on the promoter region of NFATc1. Thus, we revealed a novel role for Dpy30 in osteoclastogenesis through epigenetic mechanisms, and that it could potentially be a therapeutic target for bone destruction diseases.

Inhibition of ACLY Leads to Suppression of Osteoclast Differentiation and Function Via Regulation of Histone Acetylation

AUTHORS

Qian Guo, Honglei Kang, Jia Wang, Yimin Dong, Renpeng Peng, Hongjian Zhao, Wei Wu, Hanfeng Guan, Feng Li

ABSTRACT

ATP-citrate lyase (ACLY), generating most of the nucleocytosolic acetyl coenzyme A (acetyl-CoA) for histone acetylation, links cell metabolism to epigenetic regulation. Recent investigations demonstrated that ACLY activated by metabolic reprogramming played an essential role in both M1 and M2 macrophage activation via histone acetylation. Previous studies also revealed that histone methylation and acetylation were critical for transcriptional regulation of osteoclast-specific genes. Considering that osteoclast differentiation also undergoes metabolic reprogramming and the activity of ACLY is always Akt-dependent, we inferred that receptor activator of NF-κB (RANK) activation might enhance the activity of ACLY through downstream pathways and ACLY might play a role in osteoclast formation. In the current study, we found that ACLY was gradually activated during RANK ligand (RANKL)-induced osteoclast differentiation from bone marrow-derived macrophages (BMMs). Both ACLY knock-down and small molecular ACLY inhibitor BMS-303141 significantly decreased nucleocytosolic acetyl-CoA in BMMs and osteoclasts and suppressed osteoclast formation in vitro. BMS-303141 also suppressed osteoclast formation in vivo and prevents ovariectomy (OVX)-induced bone loss. Further investigations showed that RANKL triggered ACLY translocation into nucleus, consistent with increasing histone H3 acetylation, which was correlated to ACLY. The H3 lysine residues influenced by ACLY were in accordance with GCN5 targets. Using GCN5 knock-down and overexpression, we showed that ACLY and GCN5 functioned in the same pathway for histone H3 acetylation. Analysis of pathways downstream of RANK activation revealed that ACLY was Akt-dependent and predominately affected Akt pathway. With the help of RNA-sequencing, we discovered Rac1 as a downstream regulator of ACLY, which was involved in shACLY-mediated suppression of osteoclast differentiation, cytoskeleton organization, and signal transduction and was transcriptionally regulated by ACLY via histone H3 acetylation. To summarize, our results proved that inhibition of ATP-citrate lyase led to suppression of osteoclast differentiation and function via regulation of histone acetylation. Rac1 could be a downstream regulator of ACLY. © 2021 American Society for Bone and Mineral Research (ASBMR).